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An alkaliphilic, moderately halophilic, heterotrophic, rod-shaped, spore-forming bacterium (M30T) was isolated from a sediment sample collected from a soda lake (Lake Magadi, Tanzania). Strain M30T was strictly aerobic, catalase-positive, oxidase-negative and non-motile. Growth occurred at 12-43 °C (optimum, 25-30 °C), at pH 8.0-12 (optimum, pH 9.5-10) and at salinities of 0.5-15 % (w/v) NaCl (optimum 5 %). It utilized various sugars and organic acids as sole carbon sources and was positive for amylase, cellulase, gelatinase, protease and xylanase activities. The cell-wall peptidoglycan contained meso-diaminopimelic acid and the polar lipids consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, one unidentified lipid and one unidentified phospholipid. The DNA G+C content was 48.9 mol%. The predominant menaquinone was MK-7 and the major fatty acids (>10 %) comprised anteiso-C15 : 0, iso-C15 : 0, and anteiso-C17 : 0. Phylogenetic analysis based on 16S rRNA gene sequence affiliated M30T to the genus Bacillus and showed the highest similarities to Bacillus populi FJAT-45347T (96.4 %) and Bacillus aurantiacus K1-5T (96.2 %). Based on the data from the current polyphasic study, M30T represents a novel species of the genus Bacillus, for which the name Bacillus natronophilus sp. nov. is proposed. The type strain is M30T (=JCM 32118T=CGMCC 1.16739T=MCC 3010T).

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A Gram-stain-positive, alkaliphilic, moderately halophilic, cocci-shaped actinobacterium (strain M8T) was isolated from a sample of soda lake sediment (Lake Magadi, Tanzania). The isolate was heterotrophic, strictly aerobic, catalase-positive, oxidase-negative and formed orange-pigmented colonies in solid media. It utilized various sugars and organic acids as sole carbon sources. The organism grew at 10-38 °C, at pH 7.5-12.0 and in the presence of 1-12 % (w/v) NaCl, with optimal growth occurring at 30 °C, at pH 10 and in the presence of 5 % (w/v) NaCl. Comparative 16S rRNA gene sequence analysis showed that strain M8T belonged to the genus Nesterenkonia, sharing the closest similarities to Nesterenkoniahalobia DSM 20541T, Nesterenkoniahalophila YIM 70179T and Nesterenkoniaaethiopica DSM 17733T (97.5, 97.5 and 97.1 %, respectively). The characteristic diamino acid of strain M8T was found to be lysine and the polar lipids detected were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, two unidentified glycolipids and two unidentified phospholipids. The DNA G+C content was 61.8 mol% (genome). The strain contained MK-7, MK-9 and MK-10 as the respiratory quinones, and the major fatty acids (>10 %) comprised anteiso-C17 : 0 and anteiso-C15 : 0. On the basis of phylogenetic analyses and phenotypic data, strain M8T is considered to represent a novel species, for which the name Nesterenkonianatronophila sp. nov. is proposed. The type strain is M8T (=JCM 32100T=CGMCC 1.16706T=MCC 3367T).

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A total of 114 moderately halophilic bacteria were isolated from marine sediment environments. The isolates are belonged to 23 species based on the 16S rRNA sequence analysis. 63, 52, 47, 57, 74, 15 and 4 isolates are able to produce protease, amylase, lipase, pectinase, pulluanase, xylanase, cellulase, respectively. Combined hydrolytic enzyme activity analysis show that 15 strains present 1 hydrolytic activity, 32 strains present 2 hydrolytic activities, 21 strains present 3 hydrolytic activities, 26 strains present 4 hydrolytic activities, 11 strains present 5 hydrolytic activities and 2 strains present 6 hydrolytic activities. Hydrolase activities are widely distributed in a variety of species. The highest rates for production of protease, amylase, lipase, pectinase, pullanase, xylanase and cellulase were observed in species of B. baekryungensis, Hallobacillus sp., B. pumilus, B. megaterium or P. chungwhensis, B. amyloliquefaciens, B. pumilus, B. baekryungensis, respectively. However, the higher activities of protease, pectinase and pulluanase are frequently produced by the species of Halomonas sp. B. amyloliquefaciens or P. chungwhensis, and Vibrio sp. respectively. This investigation show that the diversity of halophilic bacteria from marine sediments could serve as a potential source of hydrolytic enzymes for industrial applications. (AU)

Um total de 114 bactérias moderadamente halofílicas foram isoladas de ambientes de sedimentos marinhos. Os isolados pertencem a 23 espécies com base na análise da sequência 16S rRNA. 63, 52, 47, 57, 74, 15 e 4 isolados são capazes de produzir protease, amilase, lipase, pectinase, pululanase, xilanase, celulase, respectivamente. A análise da atividade enzimática hidrolítica combinada mostra que 15 cepas apresentam 1 atividade hidrolítica, 32 cepas apresentam 2 atividades hidrolíticas, 21 cepas apresentam 3 atividades hidrolíticas, 26 cepas apresentam 4 atividades hidrolíticas, 11 cepas apresentam 5 atividades hidrolíticas e 2 cepas apresentam 6 atividades hidrolíticas. Atividades de hidrolase são amplamente distribuídas em uma variedade de espécies. As maiores taxas de produção de protease, amilase, lipase, pectinase, pululanase, xilanase e celulase foram observadas em espécies de B. baekryungensis, Hallobacillus sp., B. pumilus, B. megaterium ou P. chungwhensis, B. amyloliquefaciens, B. pumilus, B. baekryungensis, respectivamente. No entanto, as atividades mais elevadas de protease, pectinase e pululanase são freqüentemente produzidas pelas espécies de Halomonas sp. B. amyloliquefaciens ou P. chungwhensis e Vibrio sp. respectivamente. Esta investigação mostra que a diversidade de bactérias halofílicas de sedimentos marinhos pode servir como uma fonte potencial de enzimas hidrolíticas para aplicações industriais. (AU)

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We carried out a comparative taxonomic study of Salinivibrio proteolyticus and Salinivibrio costicola subsp. vallismortis, as well as of five halophilic strains (IB574, IB872, PR5, PR919 and PR932), isolated from salterns in Spain and Puerto Rico that were closely related to these bacteria. Multilocus sequence analysis of concatenated gyrB, recA, rpoA and rpoD housekeeping genes showed that they constituted a single cluster separate from the other species and subspecies of Salinivibrio. Experimental and in silico DNA-DNA hybridization studies indicated that they are members of the same species, with relatedness of 100-74 % and 97.8-70.0 %, respectively. The average nucleotide identity (ANI) determined for these strains was 99.7-95.6 % for ANIb and 99.7-95.7 % for OrthoANI. However, the ANI values for S. costicolasubsp.vallismortis DSM 8285T with respect to S. costicolasubsp.costicola DSM 11403T and S. costicolasubsp.alcaliphilus DSM 16359T were 78.7 and 78.9 % (ANIb) and 79.4 and 79.4 % (OrthoANI), respectively. The phylogenomic tree based on 1072 concatenated orthologous single-copy core genes confirmed that S. proteolyticus, S. costicolasubsp.vallismortis and the five new isolates constitute a coherent single phylogroup, separated from the other species and subspecies of Salinivibrio. All these data indicate that S. costicolasubsp.vallismortis is a heterotypic synonym of S. proteolyticus and we propose an emended description of this species.

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Background: DegP is a serine protease that specifically cleaves and refolds unfolding proteins in the periplasmic space of the cells. To date, there is no information regarding DegP from halophilic bacteria. Chromohalobacter salexigens BKL5 is a moderately halophilic bacterium that has the ability to grow in a media containing more than 15% salt. Therefore, the objectives of this work were to clone and overexpress DegP-encoding gene from C. salexigens BKL5 and characterize its biochemical properties. Results: DegP-encoding gene was overexpressed in Escherichia coli BL21(DE3) CodonPlus in an active form. SDS-PAGE analysis showed that the molecular weight of the recombinant DegP was 45 kDa. Size-exclusion chromatography analysis suggested that recombinant DegP was present in two multimeric states, hexameric and dodecameric, with molecular weights of 297.9 and 579.12 kDa, respectively. Both conformations were enzymatically active when casein was used as substrate for enzymatic assay. Circular dichroism analysis showed that recombinant DegP was composed of 0.21­0.29 helical content, which was comparable to the helical content in the crystal structure of E. coli DegP. The basic/acidic residue ratio of recombinant DegP was 0.56, which was slightly higher than that of DegP from extreme halophiles (average, 0.45) but significantly lower than that of DegP from nonhalophiles (average, 0.94). Conclusions: Recombinant DegP from C. salexigens BKL5 showed proteolytic activity when ß-casein was used as a substrate. In silico analysis indicated that recombinant DegP had characteristics similar to those of halophilic proteins depending on its amino acid composition.

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